Ma Lane

Changes in blood-brain barrier permeability to large and small molecules following traumatic brain injury in mice

The entry of therapeutic compounds into the brain and spinal cord is normally restricted by barrier mechanisms in cerebral blood vessels (blood–brain barrier) and choroid plexuses (blood–CSF barrier). In the injured brain, ruptured cerebral blood vessels circumvent these barrier mechanisms by allowing blood contents to escape directly into the brain parenchyma. This process may contribute to the secondary damage that follows the initial primary injury. However, this localized compromise of barrier function in the injured brain may also provide a ‘window of opportunity’ through which drugs that do not normally cross the blood–brain barriers are able to do so. This paper describes a systematic study of barrier permeability in a mouse model of traumatic brain injury using both small and large inert molecules that can be visualized or quantified. The results show that soon after trauma, both large and small molecules are able to enter the brain in and around the injury site. Barrier restriction to large (protein‐sized) molecules is restored by 4–5 h after injury. In contrast, smaller molecules (286–10 000 Da) are still able to enter the brain as long as 4 days postinjury. Thus the period of potential secondary damage from barrier disruption and the period during which therapeutic compounds have direct access to the injured brain may be longer than previously thought.

Fetuin in the Developing Neocortex of the Rat: Distribution and Origin

Immunocytochemical distribution of the fetal protein fetuin in the neocortex of developing rat brain and the presence of its mRNA, as detected by using reverse transcriptase‐polymerase chain reaction analysis, was studied in fetuses at embryonic day 15 (E15) through E22, in neonates at postnatal day 0 (P0) through P20, and in adults. Quantitative estimates of fetuin in cerebrospinal fluid (CSF) and plasma were obtained over the same period. Exogenous (bovine) fetuin injected intraperitoneally into fetal and postnatal rats was used to study the uptake of fetuin into CSF and brain and its distribution compared with endogenous fetuin; bovine albumin was used as a control. Fetuin was identified immunocytochemically in the cortical plate and subplate cells of the developing neocortex. In the rat fetus, fetuin first was apparent at E17, mainly in cell processes, but a few subplate cells also were positive. By E18, there was strong staining in subplate neurons and in inner cells of the cortical plate. At E21, these inner cells of the cortical plate were beginning to differentiate into layer VI neurons, many of which were positive for fetuin. By P0–P1, more layer VI neurons and some layer V neurons had become positive for fetuin. Fetuin immunoreactivity generally was weaker at P1, and, by P2–P3, it had disappeared from all of the layers of the developing neocortex. Bovine fetuin (but not albumin), probably taken up through CSF over the neocortical dorsal surface, had a cytoplasmic distribution; endogenous rat fetuin was both cytoplasmic and membrane bound. Thus, much of this fetuin can be accounted for by uptake, although the presence of fetuin mRNA indicates that in situ synthesis may also contribute. J. Comp. Neurol. 423:373–388, 2000.

Age‐related differences in the local cellular and molecular responses to injury in developing spinal cord of the opossum, Monodelphis domestica

Immature spinal cord, unlike adult, has an ability to repair itself following injury. Evidence for regeneration, structural repair and development of substantially normal locomotor behaviour comes from studies of marsupials due to their immaturity at birth. We have compared morphological, cellular and molecular changes in spinal cords transected at postnatal day (P)7 or P14, from 3 h to 2 weeks post‐injury, in South American opossums (Monodelphis domestica). A bridge between severed ends of cords was apparent 5 days post‐injury in P7 cords, compared to 2 weeks in P14. The volume of neurofilament (axonal) material in the bridge 2 weeks after injury was 30% of control in P7‐ but < 10% in P14‐injured cords. Granulocytes accumulated at the site of injury earlier (3 h) in P7 than in P14 (24 h)‐injured animals. Monocytes accumulated 24 h post‐injury and accumulation was greater in P14 cords. Accumulation of GFAP‐positive astrocytes at the lesion occurred earlier in P14‐injured cords. Neurites and growth cones were identified ultrastructurally in contact with astrocytes forming the bridge. Results using mouse inflammatory gene arrays showed differences in levels of expression of many TGF, TNF, cytokine, chemokine and interleukin gene families. Most of the genes identified were up‐regulated to a greater extent following injury at P7. Some changes were validated and quantified by RT‐PCR. Overall, the results suggest that at least some of the greater ability to recover from spinal cord transection at P7 compared to P14 in opossums is due to differences in inflammatory cellular and molecular responses.

Regeneration of supraspinal axons after complete transection of the thoracic spinal cord in neonatal opossums (Monodelphis domestica)

These studies define the time table and origin of supraspinal axons regenerating across a complete spinal transection in postnatal Monodelphis domestica. After lumbar (L1) spinal cord injection of fluorophore–dextran amine conjugate on postnatal (P) day 4, a consistent number of neurons could be labeled. The numbers of labeled neurons remained stable for several weeks, but subsequently declined by P60 in control animals and by P35 in animals with complete spinal transection (T4–T6) performed at P7. In control animals, 25–40% of neurons labeled with a fluorophore injected (L1) at P4 could also be double‐labeled by a second fluorophore injected (T8–T10) at different older ages. In spinally transected animals, total numbers of neurons labeled with the second marker were initially lower compared with age‐matched controls, but were not significantly different by 3 weeks after injury. The proportion of double‐labeled neurons in spinally transected animals increased from approximately 2% 1 week after injury (P14) to approximately 50% by P60, indicating that a substantial proportion of neurons with axons transected at P7 is able to regenerate and persist into adulthood. However, the proportion of axons originating from regenerating neurons made only a small contribution at older ages to total numbers of fibers growing through the injury site, because much of development of the spinal cord occurs after P7. Evidence was obtained that degenerating neurons with both apoptotic and necrotic morphologies were present in brainstem nuclei; the number of neurons with necrotic morphology was much greater in the brainstem of animals with spinal cords transected at P7. J. Comp. Neurol. 466:422–444, 2003.